Evaluation of the polyphenolic profile of native Ecuadorian stingless bee honeys (Tribe: Meliponini) and their antibiofilm activity on susceptible and multidrug-resistant pathogens: An exploratory analysis

Biofilms are associated with infections that are resistant to conventional therapies, contributing to the antimicrobial resistance crisis. The need for alternative approaches against biofilms is well-known. Although natural products like stingless bee honeys (tribe: Meliponini) constitute an alternative treatment, much is still unknown. Our main goal was to evaluate the antibiofilm activity of stingless bee honey samples against multidrug-resistant (MDR) pathogens through biomass assays, fluorescence (cell count and viability), and scanning electron (structural composition) microscopy. We analyzed thirty-five honey samples at 15% (v/v) produced by ten different stingless bee species (Cephalotrigona sp., Melipona sp., M. cramptoni, M. fuscopilosa, M. grandis, M. indecisa, M. mimetica, M. nigrifacies, Scaptotrigona problanca, and Tetragonisca angustula) from five provinces of Ecuador (Tungurahua, Pastaza, El Oro, Los Ríos, and Loja) against 24-h biofilms of Staphylococcus aureus, Klebsiella pneumoniae, Candida albicans, and Candida tropicalis. The present honey set belonged to our previous study, where the samples were collected in 2018–2019 and their physicochemical parameters, chemical composition, mineral elements, and minimal inhibitory concentration (MIC) were screened. However, the polyphenolic profile and their antibiofilm activity on susceptible and multidrug-resistant pathogens were still unknown. According to polyphenolic profile of the honey samples, significant differences were observed according to their geographical origin in terms of the qualitative profiles. The five best honey samples (OR24.1, LR34, LO40, LO48, and LO53) belonging to S. problanca, Melipona sp., and M. indecisa were selected for further analysis due to their high biomass reduction values, identification of the stingless bee specimens, and previously reported physicochemical parameters. This subset of honey samples showed a range of 63–80% biofilm inhibition through biomass assays. Fluorescence microscopy (FM) analysis evidenced statistical log reduction in the cell count of honey-treated samples in all pathogens (P <0.05), except for S. aureus ATCC 25923. Concerning cell viability, C. tropicalis, K. pneumoniae ATCC 33495, and K. pneumoniae KPC significantly decreased (P <0.01) by 21.67, 25.69, and 45.62%, respectively. Finally, scanning electron microscopy (SEM) analysis demonstrated structural biofilm disruption through cell morphological parameters (such as area, size, and form). In relation to their polyphenolic profile, medioresinol was only found in the honey of Loja, while scopoletin, kaempferol, and quercetin were only identified in honey of Los Rios, and dihydrocaffeic and dihydroxyphenylacetic acids were only detected in honey of El Oro. All the five honey samples showed dihydrocoumaroylhexose, luteolin, and kaempferol rutinoside. To the authors’ best knowledge, this is the first study to analyze stingless bees honey-treated biofilms of susceptible and/or MDR strains of S. aureus, K. pneumoniae, and Candida species.